Genome-Wide Analysis of PAPS1-Dependent Polyadenylation Identifies Novel Roles for Functionally Specialized Poly(A) Polymerases in Arabidopsis thaliana.
The poly(A) tail at 3' ends of eukaryotic mRNAs promotes their nuclear export, stability and translational efficiency, and changes in its length can strongly impact gene expression. The Arabidopsis thaliana genome encodes three canonical nuclear poly(A) polymerases, PAPS1, PAPS2 and PAPS4. As shown by their different mutant phenotypes, these three isoforms are functionally specialized, with PAPS1 modifying organ growth and suppressing a constitutive immune response. However, the molecular basis of this specialization is largely unknown. Here, we have estimated poly(A)-tail lengths on a transcriptome-wide scale in wild-type and paps1 mutants. This identified categories of genes as particularly strongly affected in paps1 mutants, including genes encoding ribosomal proteins, cell-division factors and major carbohydrate-metabolic proteins. We experimentally verified two novel functions of PAPS1 in ribosome biogenesis and redox homoeostasis that were predicted based on the analysis of poly(A)-tail length changes in paps1 mutants. When overlaying the PAPS1-dependent effects observed here with coexpression analysis based on independent microarray data, the two clusters of transcripts that are most closely coexpressed with PAPS1 show the strongest change in poly(A)-tail length and transcript abundance in paps1 mutants in our analysis. This suggests that their coexpression reflects at least partly the preferential polyadenylation of these transcripts by PAPS1 versus the other two poly(A)-polymerase isoforms. Thus, transcriptome-wide analysis of poly(A)-tail lengths identifies novel biological functions and likely target transcripts for polyadenylation by PAPS1. Data integration with large-scale co-expression data suggests that changes in the relative activities of the isoforms are used as an endogenous mechanism to co-ordinately modulate plant gene expression.


How art changes your brain: differential effects of visual art production and cognitive art evaluation on functional brain connectivity.
Visual art represents a powerful resource for mental and physical well-being. However, little is known about the underlying effects at a neural level. A critical question is whether visual art production and cognitive art evaluation may have different effects on the functional interplay of the brain's default mode network (DMN). We used fMRI to investigate the DMN of a non-clinical sample of 28 post-retirement adults (63.71 years ±3.52 SD) before (T0) and after (T1) weekly participation in two different 10-week-long art interventions. Participants were randomly assigned to groups stratified by gender and age. In the visual art production group 14 participants actively produced art in an art class. In the cognitive art evaluation group 14 participants cognitively evaluated artwork at a museum. The DMN of both groups was identified by using a seed voxel correlation analysis (SCA) in the posterior cingulated cortex (PCC/preCUN). An analysis of covariance (ANCOVA) was employed to relate fMRI data to psychological resilience which was measured with the brief German counterpart of the Resilience Scale (RS-11). We observed that the visual art production group showed greater spatial improvement in functional connectivity of PCC/preCUN to the frontal and parietal cortices from T0 to T1 than the cognitive art evaluation group. Moreover, the functional connectivity in the visual art production group was related to psychological resilience (i.e., stress resistance) at T1. Our findings are the first to demonstrate the neural effects of visual art production on psychological resilience in adulthood.


Expression of the T85A mutant of zebrafish aquaporin 3b improves post-thaw survival of cryopreserved early mammalian embryos.
While vitrification has become the method of choice for preservation of human oocytes and embryos, cryopreservation of complex tissues and of large yolk-containing cells, remains largely unsuccessful. One critical step in such instances is appropriate permeation while avoiding potentially toxic concentrations of cryoprotectants. Permeation of water and small non-charged solutes, such as those used as cryoprotectants, occurs largely through membrane channel proteins termed aquaporins (AQPs). Substitution of a Thr by an Ala residue in the pore-forming motif of the zebrafish (Dario rerio) Aqp3b paralog resulted in a mutant (DrAqp3b-T85A) that when expressed in Xenopus or porcine oocytes increased their permeability to ethylene glycol at pH 7.5 and 8.5. The main objective of this study was to test whether ectopic expression of DrAqp3b-T85A also conferred higher resistance to cryoinjury. For this, DrAqp3b-T85A + eGFP (reporter) cRNA, or eGFP cRNA alone, was microinjected into in vivo fertilized 1-cell mouse zygotes. Following culture to the 2-cell stage, appropriate membrane expression of DrAqp3b-T85A was confirmed by immunofluorescence microscopy using a primary specific antibody directed against the C-terminus of DrAqp3b. Microinjected 2-cell embryos were then cryopreserved using a fast-freezing rate and low concentration (1.5 M) of ethylene glycol in order to highlight any benefits from DrAqp3b-T85A expression. Notably, post-thaw survival rates were higher (P<0.05) for T85A-eGFP-injected than for -uninjected or eGFP-injected embryos (73±7.3 vs. 28±7.3 or 14±6.7, respectively). We propose that ectopic expression of mutant AQPs may provide an avenue to improve cryopreservation results of large cells and tissues in which current vitrification protocols yield low survival.


Conservative management for patients with chronic kidney disease refusing dialysis.
Estimates suggest that 20-30% of the deaths of patients with chronic kidney disease with indication to undergo dialysis occur after refusal to continue dialysis, discontinuation of dialysis or inability to offer dialysis on account of local conditions. Contributing factors include aging, increased comorbidity associated with chronic kidney disease, and socioeconomic status. In several occasions nephrologists will intervene, but at times general practitioners or family physicians are on their own. Knowledge of the main etiologies of chronic kidney disease and the metabolic alterations and symptoms associated to end-stage renal disease is an important element in providing patients with good palliative care. This review aimed to familiarize members of multidisciplinary care teams with the metabolic alterations and symptoms arising from chronic kidney disease treated clinically without the aid of dialysis.


Hybrid populations selectively filter gene introgression between species.
Hybrids have long been recognized as a potential pathway for gene flow between species that can have important consequences for evolution and conservation biology. However, few studies have demonstrated that genes from one species can introgress or invade another species over a broad geographic area. Using 35 genetically mapped restriction fragment length polymorphism (RFLP) markers of two species of cottonwoods (Populus fremontii x P. angustifolia) and their hybrids (n = 550 trees), we showed that the majority of the genome is prohibited from introgressing from one species into the other. However, this barrier was not absolute; Fremont cpDNA and mtDNA were found throughout the geographic range of narrowleaf cottonwood, and 20% of the nuclear markers of Fremont cottonwood introgressed varying distances (some over 100 km) into the recipient species' range. Rates of nuclear introgression were variable, but two nuclear markers introgressed as fast as the haploid, cytoplasmically inherited chloroplast and mitochondrial markers. Our genome-wide analysis provides evidence for positive, negative, and neutral effects of introgression. For example, we predict that DNA fragments that introgress through several generations of backcrossing will be small, because small fragments are less likely to contain deleterious genes. These results argue that recombination will be important, that introgression can be very selective, and that evolutionary forces within the hybrid population to effectively "filter" gene flow between species. A strong filter may make introgression adaptive, prevent genetic assimilation, lead to relaxed isolating mechanisms, and contribute to the stability of hybrid zones. Thus, rather than hybridization being a negative factor as is commonly argued, natural hybridization between native species may provide important genetic variation that impacts both ecological and evolutionary processes. Finally, we propose two hypotheses that contrast the likelihood of contemporary versus ancient introgression in this system.


High perfomance liquid chromatography in pharmaceutical analyses.
In testing the pre-sale procedure the marketing of drugs and their control in the last ten years, high performance liquid chromatography replaced numerous spectroscopic methods and gas chromatography in the quantitative and qualitative analysis. In the first period of HPLC application it was thought that it would become a complementary method of gas chromatography, however, today it has nearly completely replaced gas chromatography in pharmaceutical analysis. The application of the liquid mobile phase with the possibility of transformation of mobilized polarity during chromatography and all other modifications of mobile phase depending upon the characteristics of substance which are being tested, is a great advantage in the process of separation in comparison to other methods. The greater choice of stationary phase is the next factor which enables realization of good separation. The separation line is connected to specific and sensitive detector systems, spectrafluorimeter, diode detector, electrochemical detector as other hyphernated systems HPLC-MS and HPLC-NMR, are the basic elements on which is based such wide and effective application of the HPLC method. The purpose high performance liquid chromatography (HPLC) analysis of any drugs is to confirm the identity of a drug and provide quantitative results and also to monitor the progress of the therapy of a disease.1) Measuring presented on the Fig. 1. is chromatogram obtained for the plasma of depressed patients 12 h before oral administration of dexamethasone. It may also be used to further our understanding of the normal and disease process in the human body trough biomedical and therapeutically research during investigation before of the drugs registration. The analyses of drugs and metabolites in biological fluids, particularly plasma, serum or urine is one of the most demanding but one of the most common uses of high performance of liquid chromatography. Blood, plasma or serum contains numerous endogenous compounds often present in concentrations much greater than those of analyte. Analiyte concentrations are often low, and in the case of drugs, the endogenous compounds are sometimes structurally very similar to the drug to be measured. The binding of drugs to the plasma protein also may occur which decreases the amount of free compound that is measured. To undertake the analyses of drugs and metabolites in body fluids the analyst is facet with several problems. The first problem is due to the complex nature of the body fluid, the drugs must be isolated by an extraction technique, which ideally should provide a relatively clean extract, and the separation system must be capable of resolving the drugs of interest from co extractives. All mentioned when we are using high performance liquid chromatography require good selections of detectors, good stationary phase, eluents and adequate program during separation. UV/VIS detector is the most versatile detector used in high performance liquid chromatography it is not always ideal since it is lack of specificity means high resolution of the analyte that may be required. UV detection is preferred since it offers excellent linearity and rapid quantitative analyses can be performed against a single standard of the drug being determined. Diode array and rapid scanning detector are useful for peak identification and monitoring peak purity but they are somewhat less sensitive then single wavelength detectors. In liquid chromatography some components may have a poor UV chromophores if UV detection is being used or be completely retained on the liquid chromatography column. Fluorescence and electrochemical detector are not only considerably more sensitive towed appropriate analytes but also more selective than UV detectors for many compounds. If at all possible fluorescence detectors are sensitive, stable, selective and easy to operate. It is selectivity shows itself in the lack of frontal components observed in plasma extract whereas electrochemical detection is nearly always associated with a major frontal peak than tails considerably. To date, the most sensitive method has been the reductive electrochemical detection and giving the excellent results in the investigation on some classes of drugs. Several high performance liquid chromatography oxidative electrochemical methods have been developed for the analyses of drugs and metabolites in body fluids. Mass spectrometer as specific detector with all variation of ionisation and interface (thermo spray, moving belt etc. ) or liquid chromatography-tandem mass spectrometry2,3,4,5). NMR as selective and specific detector in high performance liquid chromatography today is also in used. The development of a non-aqueous eluent for ion-exchange separation on silica has provided an excellent system which, when used in conjugation with an electrochemical detector, permits the analyses of an extensive range of especially basic drugs and metabolites. New packing materials such as polymeric, base deactivated silica's, pyrolysed carbon and the internal surface packing should offer the improved stability and higher efficiencies for certain classes of the compounds such as basic drugs. Microbore columns should become more accepted since they offer not only improved sensitivity but also a lower solvent consumption and consequently the reduced needs to dispose of noxious solvents. Many analyses of basic drugs are still performed by the same method of the ion-exchange chromatography on unmodified silica columns with an eluent buffered to about pH 9. Neutral or weakly acidic drugs for instance barbiturates can be chromatographed on a reversed phase system whilst acidic drugs for example paracetamol, cannabis are separated either by ion suppression or ion-pair chromatography on a reversed-phase packing material. In micelar liquid chromatography micelar mobile phases in reversed-phase instead of conventional hydro organic mobile phase is used. In micelar liquid chromatography complex electrostatic hydrophobic and steric interactions exist between the solute and both stationary and mobile phases. These enable the effective separation of samples of different nature. The main advantages of the use of a micelar solution in reversed-phase liquid chromatography are the solvent and the lower cost and toxicity, the biodegradability of the solvent and the easy dissolution of analytical samples, that enables the determination of drugs in physiological fluids without the need for previous separation of the proteins present in the samples. Using tetrabutylammonium phosphate as a competing base in the investigation of sulphonamides and heptanes sulfonate as ion pairing reagent. Ion pairing reagent is term used to describe enhanced retention as the result of the addition to the mobile phase of a large ion opposite charge to the molecular ions to be separated. For molecular cations alkyl sulphates or sulfonates are generally utilised.


The remote detection of clear air turbulence by infrared radiation.
Clear air turbulence (CAT) is frequently associated with horizontal temperature gradients of the atmosphere. An instrument has been developed for detecting such gradients remotely by sensing the infrared radiation emitted by the carbon dioxide in the atmosphere. Three of these instruments have been installed on commercial jet aircraft, to evalute this technique for providing advance warning of CAT. Many light turbulence encounters have been detected as much as 80 miles (130 km) (8 min) in advance. A high false alarm rate was experienced because of temperature gradients not associated with turbulence. No severe turbulence has been encountered to date. It is hoped that the temperature effects associated with severe turbulence will be large enough to permit establishing higher alarm threshold levels which will substantially reduce the false alarm rate. Future plans include evaluation of a vertical scan mode for detection of temperature inversions.


Functionalized Thick Film Impedance Sensors for Use in In Vitro Cell Culture.
Multi-electrode arrays find application in electrophysiological recordings. The quality of the captured signals depends on the interfacial contact between electrogenic cells and the electronic system. Therefore, it requires reliable low-impedance electrodes. Low-temperature cofired ceramic technology offers a suitable platform for rapid prototyping of biological reactors and can provide both stable fluid supply and integrated bio-hardware interfaces for recordings in electrogenic cell cultures. The 3D assembly of thick film gold electrodes in in vitro bio-reactors has been demonstrated for neuronal recordings. However, especially when dimensions become small, their performance varies strongly. This work investigates the influence of different coatings on thick film gold electrodes with regard to their influence on impedance behavior. PSS layer, titanium oxynitride and laminin coatings are deposited on LTCC gold electrodes using different 2D and 3D MEA chip designs. Their impedance characteristics are compared and discussed. Titanium oxynitride layers emerged as suitable functionalization. Small 86-µm-electrodes have a serial resistance Rs of 32 kOhm and serial capacitance Cs of 4.1 pF at 1 kHz. Thick film gold electrodes with such coatings are thus qualified for signal recording in 3-dimensional in vitro cell cultures.


Bioluminescence inhibition assay for the detection of hydroxylated polychlorinated biphenyls.
Hydroxylated polychlorinated biphenyls (OH-PCBs) are an important class of contaminants that mainly originate from polychlorinated biphenyl metabolism. They may conceivably be as dangerous and persistent as the parent compounds; most prominently, OH-PCBs are endocrine disruptors. Due to increasing evidence of the presence of OH-PCBs in the environment and in living organisms, including humans, and of their toxicity, methods of detection for OH-PCBs are needed in the environmental and medical fields. Herein, we describe the development and optimization of a protein-based inhibition assay for the quantification of OH-PCBs. Specifically, the photoprotein aequorin was utilized for the detection of OH-PCBs. We hypothesized that OH-PCBs interact with aequorin, and we established that OH-PCBs actually inhibit the bioluminescence of aequorin in a dose-dependent manner. We took advantage of this phenomenon to develop an assay that is capable of detecting a wide variety of OH-PCBs with a range of detection limits, the best detection limit being 11 nM for the compound 2-hydroxy-2',3,4',5',6-pentachorobiphenyl. The viability of this system for the screening of OH-PCBs in spiked biological and environmental samples was also established. We envision the implementation of this novel bioluminescence inhibition assay as a rapid, sensitive, and cost-effective method for monitoring OH-PCBs. Furthermore, to the best of our knowledge, this is the first time aequorin has been employed to detect an analyte by the inhibition of its bioluminescence reaction. Hence, this strategy may prove to be a general approach for the development of a new generation of protein-based inhibition assays.


A harmful traditional practice in newborns with adrenocorticotropic hormone resistance syndrome: branding.
Branding refers to a traditional practice of creating 'therapeutic' burns with hot iron rods over the skin in order to treat various diseases. Although branding is a harmful practice for the body, it has been used for various illnesses including physiologic jaundice in newborns, pneumonia, and convulsions. It causes serious morbidity and delays seeking proper medical care in neonates. Innovations of modern medicine and the use of evidence-based medicine should be preferred instead of these traditional practices. We present a branded mature newborn baby who was diagnosed as having adrenocorticotropic hormone resistance syndrome. This problem is very rare in Turkey; however, it is a very important health issue and has social aspects. Therefore, this case is presented to increase awareness.


Lrrk2 pathogenic substitutions in Parkinson's disease.
Leucine-rich repeat kinase 2 (LRRK2) mutations have been implicated in autosomal dominant parkinsonism, consistent with typical levodopa-responsive Parkinson's disease. The gene maps to chromosome 12q12 and encodes a large, multifunctional protein. To identify novel LRRK2 mutations, we have sequenced 100 affected probands with family history of parkinsonism. Semiquantitative analysis was also performed in all probands to identify LRRK2 genomic multiplication or deletion. In these kindreds, referred from movement disorder clinics in many parts of Europe, Asia, and North America, parkinsonism segregates as an autosomal dominant trait. All 51 exons of the LRRK2 gene were analyzed and the frequency of all novel sequence variants was assessed within controls. The segregation of mutations with disease has been examined in larger, multiplex families. Our study identified 26 coding variants, including 15 nonsynonymous amino acid substitutions of which three affect the same codon (R1441C, R1441G, and R1441H). Seven of these coding changes seem to be pathogenic, as they segregate with disease and were not identified within controls. No multiplications or deletions were identified.


Cytochrome P450 2D6: overview and update on pharmacology, genetics, biochemistry.
Of about one dozen human P450 s that catalyze biotransformations of xenobiotics, CYP2D6 is one of the more important ones based on the number of its drug substrates. It shows a very high degree of interindividual variability, which is primarily due to the extensive genetic polymorphism that influences expression and function. This so-called debrisoquine/sparteine oxidation polymorphism has been extensively studied in many different populations and over 80 alleles and allele variants have been described. CYP2D6 protein and enzymatic activity is completely absent in less than 1% of Asian people and in up to 10% of Caucasians with two null alleles, which do not encode a functional P450 protein product. The resulting "poor metabolizer" (PM) phenotype is characterized by the inability to use CYP2D6-dependent metabolic pathways for drug elimination, which affect up to 20% of all clinically used drugs. The consequences are increased risk of adverse drug reactions or lack of therapeutic response. Today, genetic testing predicts the PM phenotype with over 99% certainty. At the other extreme, the "Ultrarapid Metabolizer" (UM) phenotype can be caused by alleles carrying multiple gene copies. "Intermediate Metabolizers" (IM) are severely deficient in their metabolism capacity compared to normal "Extensive Metabolizers" (EM), but in contrast to PMs they express a low amount of residual activity due to the presence of at least one partially deficient allele. Whereas the intricate genetics of the CYP2D6 polymorphism is becoming apparent at ever greater detail, applications in clinical practice are still rare. More clinical studies are needed to show where patients benefit from drug dose adjustment based on their genotype. Computational approaches are used to predict and rationalize substrate specificity and enzymatic properties of CYP2D6. Pharmacophore modeling of ligands and protein homology modeling are two complementary approaches that have been applied with some success. CYP2D6 is not only expressed in liver but also in the gut and in brain neurons, where endogenous substrates with high-turnover have been found. Whether and how brain functions may be influenced by polymorphic expression are interesting questions for the future.


IL-8 release and neutrophil activation by Clostridium difficile toxin-exposed human monocytes.
Neutrophil infiltration is central to the pathogenesis of Clostridium difficile toxin A-induced enterocolitis. This study examines whether monocyte activation by C. difficile toxins is instrumental in initiating neutrophil activation and recruitment. Human monocytes were exposed to low concentrations of highly purified C. difficile toxins, and the conditioned media were harvested for cytokine and functional assays. Monocytes exposed to C. difficile toxin A (10(-10) M) or toxin B (10(-12) M) released 100 and 20 times basal levels, respectively, of the neutrophil chemoattractant interleukin-8 (IL-8). Reverse transcriptase-polymerase chain reaction demonstrated a marked increase in IL-8 mRNA expression by monocytes 3 h after toxin exposure. Conditioned media from toxin A- and toxin B-treated monocytes stimulated neutrophil migration (324 and 245% of control, respectively). This effect was completely blocked by IL-8 antiserum. These media also upregulated neutrophil CD11b/CD18 and endothelial cell intercellular adhesion molecule-1 expression. C. difficile toxins, at low concentrations, potently activate monocytes to release factors, including IL-8, that facilitate neutrophil extravasation and tissue infiltration. Our findings indicate a major role for toxin-mediated monocyte and macrophage activation in C. difficile colitis.


Serum sex hormone-binding globulin and cortisol concentrations are associated with overreaching during strenuous military training.
The purpose was (a) to study the effect of an 8-week Finnish military basic training period (BT) on physical fitness, body composition, mood state, and serum biochemical parameters among new conscripts; (b) to determine the incidence of overreaching (OR); and (c) to evaluate whether initial levels or training responses differ between OR and noOR subjects. Fifty-seven males (19.7 ± 0.3 years) were evaluated before and during BT. Overreaching subjects had to fulfill 3 of 5 criteria: decreased aerobic physical fitness (VO2max), increased rating of perceived exertion (RPE) in 45-minute submaximal test at 70% of VO2max or sick absence from these tests, increased somatic or emotional symptoms of OR, and high incidence of sick absence from daily service. VO2max improved during the first 4 weeks of BT. During the second half of BT, a stagnation of increase in VO2max was observed, basal serum sex hormone-binding globulin (SHBG) increased, and insulin-like growth factor-1 and cortisol decreased. Furthermore, submaximal exercise-induced increases in cortisol, maximum heart rate, and postexercise increase in blood lactate were blunted. Of 57 subjects, 33% were classified as OR. They had higher basal SHBG before and after 4 and 7 weeks of training and higher basal serum cortisol at the end of BT than noOR subjects. In addition, in contrast to noOR, OR subjects exhibited no increase in basal testosterone/cortisol ratio but a decrease in maximal La/RPE ratio during BT. As one-third of the conscripts were overreached, training after BT should involve recovery training to prevent overtraining syndrome from developing. The results confirm that serum SHBG, cortisol, and testosterone/cortisol and maximal La/RPE ratios could be useful tools to indicate whether training is too strenuous.


Planning and process evaluation of a multi-faceted influenza vaccination implementation strategy for health care workers in acute health care settings.
Influenza transmitted by health care workers (HCWs) is a potential threat to frail patients in acute health care settings. Therefore, immunizing HCWs against influenza should receive high priority. Despite recommendations of the World Health Organization, vaccine coverage of HCWs remains low in all European countries. This study explores the use of intervention strategies and methods to improve influenza vaccination rates among HCWs in an acute care setting. The Intervention Mapping (IM) method was used to systematically develop and implement an intervention strategy aimed at changing influenza vaccination behaviour among HCWs in Dutch University Medical Centres (UMCs). Carried out during the influenza seasons 2009/2010 and 2010/2011, the interventions were then qualitatively and quantitatively evaluated by way of feedback from participating UMCs and the completion of a web-based staff questionnaire in the following spring of each season. The IM method resulted in the development of a transparent influenza vaccination intervention implementation strategy. The intervention strategy was offered to six Dutch UMCs in a randomized in a clustered Randomized Controlled Trial (RCT), where three UMCs were chosen for intervention, and three UMCs acted as controls. A further two UMCs elected to have the intervention. The qualitative process evaluation showed that HCWs at four of the five intervention UMCs were responsive to the majority of the 11 relevant behavioural determinants resulting from the needs assessment in their intervention strategy compared with only one of three control UMCs. The quantitative evaluation among a sample of HCWs revealed that of all the developed communication materials, HCWs reported the posters as the most noticeable. Our study demonstrates that it is possible to develop a structured implementation strategy for increasing the rate of influenza vaccination by HCWs in acute health care settings. The evaluation also showed that it is impossible to expose all HCWs to all intervention methods (which would have been the best case scenario). Further study is needed to (1) improve HCW exposure to intervention methods; (2) determine the effect of such interventions on vaccine uptake among HCWs; and (3) assess the impact on clinical outcomes among patients when such interventions are enacted.


Word Memory Test failure 23 times higher in mild brain injury than in parents seeking custody: the power of external incentives.
Motivation has an important influence on neuropsychological test performances. This study examined effort on the Word Memory Test (WMT) in groups with differing external incentives. 774 adults with Traumatic Brain Injury (TBI), tested as part of a Workers' Compensation, disability or personal injury claim stood to gain financially by appearing impaired on testing. In contrast, parents ordered by the Court to undergo a parenting assessment were highly motivated to do their best on cognitive tests because their goal was to regain custody of their children. Consistent with these assumptions, 98.3% of 118 parents seeking child custody passed the WMT effort subtests but in cases of mild TBI the pass rate on the WMT was only 60%. The WMT failure rate in the mild TBI sample was 23 times higher than in the group of parents seeking custody. WMT failure was twice as frequent in the mild TBI group than in those with more severe TBI. WMT failure was also much higher in adults with mild TBI than in children with significant impairment from various clinical conditions. Such differences in failure rates on the WMT effort subtests cannot be explained by differences in cognitive skills but they are explainable by differences in external incentives. The findings support the recommendation that objective tests of effort should be used when evaluating cognitive impairment.


Diffusion tensor imaging correlates of cognitive-motor decline in normal aging and increased Alzheimer's disease risk.
Alzheimer's disease (AD) is typically associated with impairments in memory and other aspects of cognition, while deficits in complex movements are commonly observed later in the course of the disease. Recent studies, however, have indicated that subtle deteriorations in visuomotor control under cognitively demanding conditions may in fact be an early identifying feature of AD. Our previous work has shown that the ability to perform visuomotor tasks that rely on visual-spatial and rule-based transformations is disrupted in prodromal and preclinical AD. Here, in a sample of 30 female participants (10 young: mean age = 26.6 ± 2.7, 10 low AD risk: mean age = 58.7 ± 5.6, and 10 high AD risk: mean age = 58.5 ± 6.9), we test the hypothesis that these cognitive-motor impairments are associated with early AD-related brain alterations. Using diffusion-weighted magnetic resonance imaging, we examined changes in white matter (WM) integrity associated with normal aging and increased AD risk, and assessed the relationship between these underlying WM alterations and cognitive-motor performance. Our whole-brain analysis revealed significant age-related declines in WM integrity, which were more widespread in high relative to low AD risk participants. Furthermore, analysis of mean diffusivity measures within isolated WM clusters revealed a stepwise decline in WM integrity across young, low AD risk, and high AD risk groups. In support of our hypothesis, we also observed that lower WM integrity was associated with poorer cognitive-motor performance. These results are the first to demonstrate a relationship between AD-related WM alterations and impaired cognitive-motor control. The application of these findings may provide a novel clinical strategy for the early detection of individuals at increased AD risk.


Meta-analyses in orthopaedic surgery. A systematic review of their methodologies.
The number and quality of well-designed scientific studies in the orthopaedic literature are limited. The purpose of this review was to determine the methodological qualities of published meta-analyses on orthopaedic-surgery-related topics. A systematic review of meta-analyses was conducted. A search of the Medline database provided lists of meta-analyses in orthopaedics published from 1969 to 1999. Extensive manual searches of major orthopaedic journals, bibliographies of major orthopaedic texts, and personal files identified additional studies. Of 601 studies identified, forty met the criteria for eligibility. Two investigators each assessed the quality of the studies under blinded conditions, and they abstracted relevant data. More than 50% of the meta-analyses included in this review were published after 1994. We found that 88% had methodological flaws that could limit their validity. The main deficiency was a lack of information on the methods used to retrieve and assess the validity of the primary studies. Regression analysis revealed that meta-analyses authored in affiliation with an epidemiology department and those published in nonsurgical journals were associated with higher scores for quality. Meta-analyses with lower scores for quality tended to report positive findings. The meta-analyses that focused upon fracture treatment and degenerative disease (hip, knee, or spine) had significantly lower mean quality scores than did meta-analyses that examined thrombosis prevention and diagnostic tests (p < 0.05). The majority of meta-analyses on orthopaedic-surgery-related topics have methodological limitations. Limitation of bias and improvement in the validity of the meta-analyses can be achieved by adherence to strict scientific methodology. However, the ultimate quality of a meta-analysis depends on the quality of the primary studies on which it is based. A meta-analysis is most persuasive when data from high-quality randomized trials are pooled.


Cell contact accelerates replicative senescence of human mesenchymal stem cells independent of telomere shortening and p53 activation: roles of Ras and oxidative stress.
Mesenchymal stem cells (MSCs) are of great therapeutic potentials due to their multilineage differentiation capabilities. Before transplantation, in vitro culture expansion of MSCs is necessary to get desired cell number. We observed that cell contact accelerated replicative senescence during such process. To confirm the finding as well as to investigate the underlying mechanisms, we cultured both human bone marrow- and umbilical cord blood-derived MSCs under noncontact culture (subculture performed at 60-70% of confluence), or contact culture (cell passage performed at 100% of confluence). It was found that MSCs reached cellular senescence earlier in contact culture, and the doubling time was significantly prolonged. Marked increase of senescence-associated β-galactosidase-positive staining was also observed as a result of cell contact. Cell cycle analysis revealed increased frequency of cell cycle arrest after contact culture. It was noted, however, that the telomere length was not altered during contact-induced acceleration of senescence. Moreover, cell cycle checkpoint regulator P53 expression was not affected by cell contact. Marked increase in intracellular reactive oxygen species (ROS) and a concomitant decrease in the activities of antioxidative enzymes were also observed during contact-induced senescence. Importantly, increased p16(INK4a) following Ras upregulation was found after contact culture. Taken together, cell contact induced accelerated senescence of MSCs, which is telomere shortening and p53 independent. ROS accumulation due to defective ROS clearance function together with Ras and p16(INK4a) upregulation play an important role in contact-induced senescence of MSCs. Overconfluence should therefore be avoided during in vitro culture expansion of MSCs in order to maintain their qualities for clinical application purposes. The contact-induced senescence model reported in this study will serve as a useful model system that allows further study of the molecular mechanisms of senescence in MSCs.


Predictors of exclusive breastfeeding at least 8 weeks among Asian and Native Hawaiian or other Pacific Islander race subgroups in Hawaii, 2004-2008.
Breastfeeding is nurturing, cost-effective, and beneficial for the health of mother and child. Babies receiving formula are sick more often and are at higher risk for childhood obesity, diabetes, asthma, and other conditions compared with breastfed children. National and international organizations recommend exclusive breastfeeding for 6 months. Exclusive breastfeeding in Asian and Native Hawaiian or Other Pacific Islander (NHOPI) subgroups is not well characterized. Data from the 2004-2008 Hawaii Pregnancy Risk Assessment Monitoring System, a population-based surveillance system on maternal behaviors and experiences before, during, and after pregnancy, were analyzed for 8,508 mothers with a recent live birth. We examined exclusive breastfeeding status for at least 8 weeks. We calculated prevalence risk ratios across maternal race groups accounting for maternal and socio-demographic characteristics. The overall estimate of exclusive breastfeeding for at least 8 weeks was 36.3%. After adjusting for maternal age, pre-pregnancy weight, cesarean delivery, return to work/school, and self-reported postpartum depressive symptoms, the racial differences in prevalence ratios for exclusive breastfeeding for each ethnic group compared to Whites were: Samoan (aPR = 0.54; 95% CI 0.43-0.69), Filipino (aPR = 0.58; 95% CI 0.53-0.63), Japanese (aPR = 0.58; 95% CI 0.52-0.65), Chinese (aPR = 0.64; 95% CI 0.58-0.70), Native Hawaiian (aPR = 0.67; 95% CI 0.61-0.72), Korean (aPR = 0.72; 95% CI 0.64-0.82), and Black (aPR = 0.79; 95% CI 0.65-0.96) compared to white mothers. Providers and community groups should be aware that just over one-third of mothers breastfeed exclusively at least 8 weeks with lower rates among Asian, NHOPI, and Black mothers. Culturally appropriate efforts to promote exclusive breastfeeding are recommended particularly among Asian subgroups that have high breastfeeding initiation rates that do not translate into high exclusivity rates.


Obesity is associated with activated and insulin resistant immune cells.
Obesity and type 2 diabetes mellitus are characterized by insulin resistance and 'low-grade inflammation'; however, the pathophysiological link is poorly understood. To determine the relative contribution of obesity and insulin resistance to systemic 'inflammation', this study comprehensively characterized circulating immune cells in different grades of obesity. Immune cell phenotypes and activation status were analysed by flow cytometry cross-sectionally in morbidly obese (n = 16, body mass index (BMI) 42.2 ± 5.4 kg/m2), overweight (n = 13, BMI 27.4 ± 1.6 kg/m2) and normal weight (n = 12, BMI 22.5 ± 1.9 kg/m2) subjects. Obese, but not overweight subjects, had increased activation marker expression on neutrophils, monocytes, T-lymphocytes and polarization of T helper cells towards a pro-inflammatory type 1-phenotype (Th1). Th1 numbers correlated positively with the degree of insulin resistance (homeostasis model assessment, p < 0.05). Lymphocytes from obese subjects showed reduced insulin-stimulated AKT-phosphorylation in vitro. Supra-physiological insulin concentrations did not affect T-cell differentiation, which under normal circumstances would promote an anti-inflammatory T helper type 2-phenotype. These results show that morbid obesity is characterized by circulating immune cells that are activated and insulin resistant, with the T-cell balance polarized towards a pro-inflammatory Th1 phenotype. The loss of insulin-induced suppression of inflammatory phenotypes in circulating immune cells could contribute to the systemic and adipose tissue inflammation found in morbid obesity.


Sensitivity of the discrete-time Kaplan-Meier estimate to nonignorable censoring: Application in a clinical trial.
Untestable assumptions about the association between survival and censoring times can affect the validity of estimates of the survival distribution including the Kaplan-Meier (KM) nonparametric maximum likelihood estimate (MLE). This paper explores the sensitivity of the KM curve to nonignorable censoring by extending the index of local sensitivity to nonignorability (ISNI; Troxel et al., Statistica Sinica, 14, 1221-1237, 2004; Zhang and Heitjan, Biometrics, 62, 1260-1268, 2006) to the case of a nonparametric survival model. The method involves, first, specifying a coarse-data selection model to describe the association between the failure and censoring processes and then evaluating the slope of the nonparametric survival MLE ordinate with respect to a nonignorability parameter in the neighborhood of the ignorable model. We define the nonparametric MLE of the survival curve for a fixed value of the nonignorability parameter and show in a simulation that ISNI analysis effectively captures local sensitivity to nonignorability. The method measures sensitivity in the sense of identifying functionals of the nonparametric MLE that nonignorability, if present, can affect substantially. We demonstrate the method with an application to a trial comparing mechanical assistance to optimal medical management in the treatment of end-stage heart failure.


Revision after osteotomy for gonarthrosis. A 10-19-year follow-up of 314 cases.
During a 10-year period, 314 tibial osteotomies were performed for medial gonarthrosis. At follow-up after 10-19 years, 62 knees had been revised by arthroplasty (52 cases) or reosteotomy (10 cases). The revision rate was 54/170 in undercorrected knees and 8/144 in knees with normalization or overcorrection of the hip-knee-ankle angle. Our results confirm that tibial osteotomy for gonarthrosis requires proper indications and precise surgery. Perhaps, given another 10 years, our osteotomies will have outlived contemporary arthroplasties.


Genome-wide and molecular evolution analysis of the subtilase gene family in Vitis vinifera.
Vitis vinifera (grape) is one of the most economically significant fruit crops in the world. The availability of the recently released grape genome sequence offers an opportunity to identify and analyze some important gene families in this species. Subtilases are a group of subtilisin-like serine proteases that are involved in many biological processes in plants. However, no comprehensive study incorporating phylogeny, chromosomal location and gene duplication, gene organization, functional divergence, selective pressure and expression profiling has been reported so far for the grape. In the present study, a comprehensive analysis of the subtilase gene family in V. vinifera was performed. Eighty subtilase genes were identified. Phylogenetic analyses indicated that these subtilase genes comprised eight groups. The gene organization is considerably conserved among the groups. Distribution of the subtilase genes is non-random across the chromosomes. A high proportion of these genes are preferentially clustered, indicating that tandem duplications may have contributed significantly to the expansion of the subtilase gene family. Analyses of divergence and adaptive evolution show that while purifying selection may have been the main force driving the evolution of grape subtilases, some of the critical sites responsible for the divergence may have been under positive selection. Further analyses of real-time PCR data suggested that many subtilase genes might be important in the stress response and functional development of plants. Tandem duplications as well as purifying and positive selections have contributed to the functional divergence of subtilase genes in V. vinifera. The data may contribute to a better understanding of the grape subtilase gene family.


China's water environment at the beginning of the 21st century: challenges and countermeasures.
Water environment crisis is a common challenge throughout the world, and the situation is becoming worse. This paper analyzes the development trend of water environment worldwide, and then evaluates the status of China's water environment and the progress in the work of protection of the water environment. According to the future development of economy and society in China, the paper discusses the severe challenges in water environment that China faces at the beginning of the 21st century and proposes related countermeasures.


Hybrid-pumped, gain-switched 1120 nm ytterbium-doped fiber laser.
A monolithic, hybrid-pumped, gain-switched, single-mode, 1120 nm Yb-doped fiber laser is reported. Pumped by a 1064 nm pulse fiber source and a continuous-wave 976 nm laser diode, a record output pulse energy of 91.7 μJ at repetition rate of 10 kHz is obtained. The full width at half-maximum of the pulses is 4.1 μs, and the slope efficiency corresponding to the 1064 nm laser is more than 46%. The pulse build-up time and a pulsewidth decrease with an increase in the pump pulse energy are observed. In the gain-switched pulses, mode-lock-resembling pulses are detected. The subpulse's duration is about 415 ns, including the cavity round-trip time, and the width varied from 80 to 320 ns, which is longer than the traditional mode-locked laser.


Post-traumatic stress disorder among recently diagnosed patients with HIV/AIDS in South Africa.
This study examined the prevalence of and factors associated with post-traumatic stress disorder in recently diagnosed HIV/AIDS patients in South Africa. One hundred and forty-nine (44 male, 105 female) recently diagnosed HIV/AIDS patients (mean duration since diagnosis = 5.8 months, SD = 4.1) were evaluated. Subjects were assessed using the MINI International Neuropsychiatric Interview (MINI), the Carver Brief COPE coping scale and the Sheehan Disability Scale. In addition, previous exposures to trauma and past risk behaviours were assessed. Twenty-two patients (14.8%) met criteria for PTSD. Current psychiatric conditions more likely to be associated with PTSD included major depressive disorder (29% in PTSD patients versus 7% in non-PTSD patients, p = 0.004), suicidality (54% versus 11%, p = 0.001) and social anxiety disorder (40% versus 13%, p = 0.04). Further patients with PTSD reported significantly more work impairment and demonstrated a trend towards higher usage of alcohol as a means of coping. Discriminant function analysis indicated that female gender and a history of sexual violation in the past year were significantly associated with a diagnosis of PTSD. Patients whose PTSD was a direct result of an HIV/AIDS diagnosis (8/22) did not differ from other patients with PTSD on demographic or clinical features. In the South African context, PTSD is not an uncommon disorder in patients with HIV/AIDS. In some cases, PTSD is secondary to the diagnosis of HIV/AIDS but in most cases it is seen after other traumas, with sexual violation and intimate partner violence in women being particularly important.


Increased phosphorylation of the RcsB regulator of the RcsCDB phosphorelay in strains of Dickeya dadantii devoid of osmoregulated periplasmic glucans revealed by Phos-tag gel analysis.
Osmoregulated periplasmic glucans (OPGs) are general constituents of many proteobacteria. OPGs are important factors required for full virulence in many pathogens including Dickeya dadantii. D. dadantii causes the soft-rot disease in a wide range of plant species. The pleiotropic phenotype of opg-negative strains includes total loss of virulence and motility, and is linked to the constitutive activation of the RcsCDB phosphorelay, deduced from expression analysis of genes of the RcsCDB regulon. The constitutive activation of the RcsCDB phosphorelay in an opg-negative strain was demonstrated by direct analysis of the phosphorylation level of the RcsB regulator protein in vivo by using a Phos-tag retardation gel approach, and was correlated with the phenotype and the expression of motility genes. Data revealed a low level of RcsB phosphorylated form in the wild-type strain and a slight increase of phosphorylation in opgG mutant strains sufficient to induce the pleiotropic phenotype observed.


Genetic scrambling as a defence against meiotic drive.
Genetic recombination has important consequences, including the familiar rules of Mendelian genetics. Here we present a new argument for the evolutionary function of recombination based on the hypothesis that meiotic drive systems continually arise to threaten the fairness of meiosis. These drive systems act at the expense of the fitness of the organism as a whole for the benefit of the genes involved. We show that genes increasing crossing over are favoured, in the process of breaking up drive systems and reducing the fitness loss to organisms.


Hierarchical cultural values predict success and mortality in high-stakes teams.
Functional accounts of hierarchy propose that hierarchy increases group coordination and reduces conflict. In contrast, dysfunctional accounts claim that hierarchy impairs performance by preventing low-ranking team members from voicing their potentially valuable perspectives and insights. The current research presents evidence for both the functional and dysfunctional accounts of hierarchy within the same dataset. Specifically, we offer empirical evidence that hierarchical cultural values affect the outcomes of teams in high-stakes environments through group processes. Experimental data from a sample of expert mountain climbers from 27 countries confirmed that climbers expect that a hierarchical culture leads to improved team coordination among climbing teams, but impaired psychological safety and information sharing compared with an egalitarian culture. An archival analysis of 30,625 Himalayan mountain climbers from 56 countries on 5,104 expeditions found that hierarchy both elevated and killed in the Himalayas: Expeditions from more hierarchical countries had more climbers reach the summit, but also more climbers die along the way. Importantly, we established the role of group processes by showing that these effects occurred only for group, but not solo, expeditions. These findings were robust to controlling for environmental factors, risk preferences, expedition-level characteristics, country-level characteristics, and other cultural values. Overall, this research demonstrates that endorsing cultural values related to hierarchy can simultaneously improve and undermine group performance.


Delayed Diagnosis of Pelvic Hematoma without Fracture Due to Military Parachuting.
The U.S. military has been conducting static-line parachute jumps for nearly a century. Beginning with World War II, military forces have also employed full-scale airborne operations as a method for insertion into combat. Through the years, injuries from blunt trauma as a result of static-line parachute jumps have evolved little with the refinement of equipment, training, and tactics. Parachute jumps continue to invoke primarily musculoskeletal injuries, especially to the lower extremities, back, neck, and head. These injuries are usually straightforward in their presentation and diagnosis. We describe the delayed diagnosis of a pelvic hematoma due to an uncommon blunt trauma jump injury. The purpose of this case report is to increase awareness of injury patterns during paratrooper operations, as well as to review the diagnosis and management of occult hemorrhage. Specific objectives for the readers are to (1) know the common injury types and patterns for airborne operations, (2) know the descent rate of T-10C/D parachutes and factors influencing the rate, (3) recognize signs and symptoms associated with a pelvic hematoma, and (4) recognize common complications resulting from a pelvic hematoma.


Prefix Stripping Re-Re-Revisited: MEG Investigations of Morphological Decomposition and Recomposition.
We revisit a long-standing question in the psycholinguistic and neurolinguistic literature on comprehending morphologically complex words: are prefixes and suffixes processed using the same cognitive mechanisms? Recent work using Magnetoencephalography (MEG) to uncover the dynamic temporal and spatial responses evoked by visually presented complex suffixed single words provide us with a comprehensive picture of morphological processing in the brain, from early, form-based decomposition, through lexical access, grammatically constrained recomposition, and semantic interpretation. In the present study, we find that MEG responses to prefixed words reveal interesting early differences in the lateralization of the form-based decomposition response compared to the effects reported in the literature for suffixed words, but a very similar post-decomposition profile. These results not only address a question stretching back to the earliest days of modern psycholinguistics, but also add critical support and nuance to our much newer emerging understanding of spatial organization and temporal dynamics of morphological processing in the human brain.


Omission of adjuvant therapy after gastric cancer resection: development of a validated risk model.
NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines) for Gastric Cancer recommend adjuvant chemotherapy with or without radiotherapy following after resection of gastric adenocarcinoma (GA) for patients who have not received neoadjuvant therapy. Despite frequent noncompliance with NCCN Guidelines nationally, risk factors underlying adjuvant therapy omission (ATom) have not been well characterized. We developed an internally validated preoperative instrument stratifying patients by incremental risk of ATom. The National Cancer Data Base was queried for patients with stage IB-III GA undergoing gastrectomy; those receiving neoadjuvant therapy were excluded. Multivariable models identified factors associated with ATom between 2006 and 2011. Internal validation was performed using bootstrap analysis; model discrimination and calibration were assessed using k-fold cross-validation and Hosmer-Lemeshow procedures, respectively. Using weighted β-coefficients, a simplified Omission Risk Score (ORS) was created to stratify ATom risk. The impact of ATom on overall survival (OS) was examined in ORS risk-stratified cohorts. In 4,728 patients (median age, 70 years; 64.8% male), 53.7% had ATom. The bootstrap-validated model identified advancing age, comorbidity, underinsured/uninsured status, proximal tumor location, and clinical T1/2 and N0 tumors as independent ATom predictors, demonstrating good discrimination. The simplified ORS, stratifying patients into low-, moderate-, and high-risk categories, predicted incremental risk of ATom (30% vs 53% vs 80%, respectively) and progressive delay to adjuvant therapy initiation (median time, 51 vs 55 vs 61 days, respectively). Patients at moderate/high-risk of ATom demonstrated worsening risk-adjusted mortality compared with low-risk patients (median OS, 26.4 vs 29.2 months). This ORS may aid in rational selection of multimodality treatment sequence in GA.


Trimellitic anhydride-induced allergic response in the lung: role of the complement system in cellular changes.
Trimellitic anhydride (TMA) is a small molecular weight industrial compound that will cause asthma-like symptoms in humans. Some of these TMA-induced symptoms can be reproduced in the guinea pig. In the guinea pig model of TMA-induced asthma, intratracheal instillation of TMA coupled to guinea pig serum albumin causes an immediate bronchoconstriction and increase in airway microvascular leakage with concomitant decrease in circulating platelets and white blood cells and subsequent cellular infiltration of mononuclear cells, neutrophils and eosinophils into the bronchoalveolar lavage fluid. In addition, in the lung tissue an increase in eosinophil peroxidase activity (a measure of eosinophil numbers) occurs. The purpose of this study was to determine whether complement system activation was essential for any of these TMA-induced events. Guinea pigs pretreated with cobra venom factor (CVF) had significantly reduced amounts of complement component C3 in the lavage fluid 24 hours after TMA conjugated to guinea pig serum albumin challenge indicating that the CVF treatment was successful in depleting complement proteins. Pretreatment with CVF did not affect the immediate TMA-induced bronchoconstriction nor the TMA-induced microvascular leakage. In animals depleted of the complement system by pretreatment with CVF the TMA-induced increase in mononuclear cells, total white blood cells, red blood cells, and EPO activity in the bronchoalveolar lavage was significantly reduced. Thus, our results suggest that in the guinea pig, the complement system is an important source of mediators for cellular infiltration into the lung after exposure to this acid anhydride and that inhibiting complement activation may be useful in preventing the inflammatory cell infiltration in TMA-induced asthma.


Identification of Global and Ligand-Specific Calcium Sensing Receptor Activation Mechanisms.
Calcium sensing receptor (CaSR) positive allosteric modulators (PAMs) are therapeutically important. However, few are approved for clinical use, in part due to complexities in assessing allostery at a receptor where the endogenous agonist (extracellular calcium) is present in all biologic fluids. Such complexity impedes efforts to quantify and optimize allosteric drug parameters (affinity, cooperativity, and efficacy) that dictate PAM structure-activity relationships (SARs). Furthermore, an underappreciation of the structural mechanisms underlying CaSR activation hinders predictions of how PAM SAR relates to in vitro and in vivo activity. Herein, we combined site-directed mutagenesis and calcium mobilization assays with analytical pharmacology to compare modes of PAM binding, positive modulation, and agonism. We demonstrate that 3-(2-chlorophenyl)-N-((1R)-1-(3-methoxyphenyl)ethyl)-1-propanamine (NPS R568) binds to a 7 transmembrane domain (7TM) cavity common to class C G protein-coupled receptors and used by (αR)-(-)-α-methyl-N-[3-[3-[trifluoromethylphenyl]propyl]-1-napthalenemethanamine (cinacalcet) and 1-benzothiazol-2-yl-1-(2,4-dimethylphenyl)-ethanol (AC265347); however, there are subtle distinctions in the contribution of select residues to the binding and transmission of cooperativity by PAMs. Furthermore, we reveal some common activation mechanisms used by different CaSR activators, but also demonstrate some differential contributions of residues within the 7TM bundle and extracellular loops to the efficacy of the PAM-agonist, AC265347, versus cooperativity. Finally, we show that PAMS potentiate the affinity of divalent cations. Our results support the existence of both global and ligand-specific CaSR activation mechanisms and reveal that allosteric agonism is mediated in part via distinct mechanisms to positive modulation.


Multidimensional analysis of the insoluble sub-proteome of Oceanobacillus iheyensis HTE831, an alkaliphilic and halotolerant deep-sea bacterium isolated from the Iheya ridge.
We report the first proteomic analysis of the insoluble sub-proteome of the alkaliphilic and halotolerant deep-sea bacterium Oceanobacillus iheyensis HTE831. A multidimensional gel-based and gel-free analysis was utilised and a total of 4352 peptides were initially identified by automated MS/MS identification software. Automated curation of this list using PROVALT reduced our peptide list to 467 uniquely identified peptides that resulted in the positive identification of 153 proteins. These identified proteins were functionally classified and physiochemically characterised. Of 26 proteins identified as hypothetical conserved, we have assigned function to all but four. A total of 41 proteins were predicted to possess signal peptides. In silico investigation of these proteins allowed us to identify three of the five bacterial classes of signal peptide, namely: (i) twin-arginine translocation; (ii) Sec-type and (iii) lipoprotein transport. Our proteomic strategy has also allowed us to identify, at neutral pH, a number of proteins described previously as belonging to two putative transport systems believed to be of importance in the alkaliphilic adaptation of O. iheyensis HTE831.


Predicting foot ulcers in patients with diabetes: a systematic review and meta-analysis.
Clinical guidelines recommend that all patients with diabetes should be screened annually to establish their risk of foot ulceration. The aim of this systematic review was to quantify the predictive value of diagnostic tests, physical signs and elements from the patient's history in relation to diabetic foot ulcers. Observational studies were identified from: electronic databases (MEDLINE, EMBASE and CINAHL); bibliographies of studies meeting the inclusion criteria; review articles and clinical guidelines; direct contact with authors. Published reports of cohort and case-control studies were considered for inclusion. Pooled estimates were calculated from absolute numbers as weighted mean differences, standard mean differences or odds ratios. Adjusted odds ratios from published reports were also extracted. We identified five case-control and 11 cohort studies. The incidence of foot ulcers ranged from 8% to 17% in the cohort studies, with varying lengths of follow-up. Diagnostic tests and physical signs that detect peripheral neuropathy (biothesiometry, monofilaments and absent ankle reflexes), and those that detect excessive plantar pressure (peak plantar pressure and joint deformity) were all significantly associated with future diabetic foot ulceration. However, there was a paucity of evidence concerning the predictive value of symptoms and signs. Further research is needed to establish the independent factors associated with diabetic foot ulceration, particularly elements from a patient's history and physical examination.


Prognostic value of estimating functional capacity with the use of the duke activity status index in stable patients with chronic heart failure.
Over the years, several methods have been developed to reliably quantify functional capacity in patients with heart failure. Few studies have investigated the prognostic value of these assessment tools beyond cardiorenal prognostic biomarkers in stable patients with chronic heart failure. We administered the Duke Activity Status Index (DASI) questionnaire, a self-assessment tool comprising 12 questions for estimating functional capacity, to 1,700 stable nonacute coronary syndrome patients with history of heart failure who underwent elective diagnostic coronary angiography with 5-year follow-up of all-cause mortality. In a subset of patients (n = 800), B-type natriuretic peptide (BNP) was measured. In our study cohort, the median DASI score was 26.2 (interquartile range [IQR] 15.5-42.7). Low DASI score provided independent prediction of a 3.3-fold increase in 5-year mortality risk (quartile 1 vs quartile 4: hazard ratio [HR] 3.33, 95% confidence interval [CI] 2.57-4.36; P < .0001). After adjusting for traditional risk factors, BNP, and estimated glomerular filtration rate, low DASI score still conferred a 2.6-fold increase in mortality risk (HR 2.57, 95% CI 1.64-4.15; P < .0001). A simple self-assessment tool of functional capacity provides independent and incremental prognostic value for mortality prediction in stable patients with chronic heart failure beyond cardiorenal biomarkers.


4D cryo-electron microscopy of proteins.
Cryo-electron microscopy is a form of transmission electron microscopy that has been used to determine the 3D structure of biological specimens in the hydrated state and with high resolution. We report the development of 4D cryo-electron microscopy by integrating the fourth dimension, time, into this powerful technique. From time-resolved diffraction of amyloid fibrils in a thin layer of vitrified water at cryogenic temperatures, we were able to detect picometer movements of protein molecules on a nanosecond time scale. Potential future applications of 4D cryo-electron microscopy are numerous, and some are discussed here.


Styrene and styrene oxide concentrations in the air during the lamination process in the reinforced plastics industry.
Styrene and styrene oxide concentrations were measured during the lamination process in the reinforced plastics industry. The mean concentration of styrene in the personal samples was 130 ppm, the highest value measured being 350 ppm. The average concentration for styrene oxide alone was 0.1 ppm, whereas the corresponding measurement for styrene oxide and its decomposition products combined was 0.7 ppm. In comparison then, the concentrations of styrene oxide and its derivatives were much lower (about 0.5% of the total) than those of styrene.


Metal ion and ligand binding of integrin α5β1.
Integrin α5β1 binds to an Arg-Gly-Asp (RGD) motif in its ligand fibronectin. We report high-resolution crystal structures of a four-domain α5β1 headpiece fragment, alone or with RGD peptides soaked into crystals, and RGD peptide affinity measurements. The headpiece crystallizes in a closed conformation essentially identical to that seen previously for α5β1 complexed with a Fab that allosterically inhibits ligand binding by stabilizing the closed conformation. Soaking experiments show that binding of cyclic RGD peptide with 20-fold higher affinity than a linear RGD peptide induces conformational change in the β1-subunit βI domain to a state that is intermediate between closed (low affinity) and open (high affinity). In contrast, binding of a linear RGD peptide induces no shape shifting. However, linear peptide binding induces shape shifting when Ca(2+) is depleted during soaking. Ca(2+) bound to the adjacent to metal ion-dependent adhesion site (ADMIDAS), at the locus of shape shifting, moves and decreases in occupancy, correlating with an increase in affinity for RGD measured when Ca(2+) is depleted. The results directly demonstrate that Ca(2+) binding to the ADMIDAS stabilizes integrins in the low-affinity, closed conformation. Comparisons in affinity between four-domain and six-domain headpiece constructs suggest that flexible integrin leg domains contribute to conformational equilibria. High-resolution views of the hybrid domain interface with the plexin-semaphorin-integrin (PSI) domain in different orientations show a ball-and-socket joint with a hybrid domain Arg side chain that rocks in a PSI domain socket lined with carbonyl oxygens.


Unorthodox PCNA Binding by Chromatin Assembly Factor 1.
The eukaryotic DNA replication fork is a hub of enzymes that continuously act to synthesize DNA, propagate DNA methylation and other epigenetic marks, perform quality control, repair nascent DNA, and package this DNA into chromatin. Many of the enzymes involved in these spatiotemporally correlated processes perform their functions by binding to proliferating cell nuclear antigen (PCNA). A long-standing question has been how the plethora of PCNA-binding enzymes exert their activities without interfering with each other. As a first step towards deciphering this complex regulation, we studied how Chromatin Assembly Factor 1 (CAF-1) binds to PCNA. We demonstrate that CAF-1 binds to PCNA in a heretofore uncharacterized manner that depends upon a cation-pi (π) interaction. An arginine residue, conserved among CAF-1 homologs but absent from other PCNA-binding proteins, inserts into the hydrophobic pocket normally occupied by proteins that contain canonical PCNA interaction peptides (PIPs). Mutation of this arginine disrupts the ability of CAF-1 to bind PCNA and to assemble chromatin. The PIP of the CAF-1 p150 subunit resides at the extreme C-terminus of an apparent long α-helix (119 amino acids) that has been reported to bind DNA. The length of that helix and the presence of a PIP at the C-terminus are evolutionarily conserved among numerous species, ranging from yeast to humans. This arrangement of a very long DNA-binding coiled-coil that terminates in PIPs may serve to coordinate DNA and PCNA binding by CAF-1.


AVATREE: An open-source computational modelling framework modelling Anatomically Valid Airway TREE conformations.
This paper presents AVATREE, a computational modelling framework that generates Anatomically Valid Airway tree conformations and provides capabilities for simulation of broncho-constriction apparent in obstructive pulmonary conditions. Such conformations are obtained from the personalized 3D geometry generated from computed tomography (CT) data through image segmentation. The patient-specific representation of the bronchial tree structure is extended beyond the visible airway generation depth using a knowledge-based technique built from morphometric studies. Additional functionalities of AVATREE include visualization of spatial probability maps for the airway generations projected on the CT imaging data, and visualization of the airway tree based on local structure properties. Furthermore, the proposed toolbox supports the simulation of broncho-constriction apparent in pulmonary diseases, such as chronic obstructive pulmonary disease (COPD) and asthma. AVATREE is provided as an open-source toolbox in C++ and is supported by a graphical user interface integrating the modelling functionalities. It can be exploited in studies of gas flow, gas mixing, ventilation patterns and particle deposition in the pulmonary system, with the aim to improve clinical decision making.


Using Electronic Health Record Data to Rapidly Identify Children with Glomerular Disease for Clinical Research.
The rarity of pediatric glomerular disease makes it difficult to identify sufficient numbers of participants for clinical trials. This leaves limited data to guide improvements in care for these patients. The authors developed and tested an electronic health record (EHR) algorithm to identify children with glomerular disease. We used EHR data from 231 patients with glomerular disorders at a single center to develop a computerized algorithm comprising diagnosis, kidney biopsy, and transplant procedure codes. The algorithm was tested using PEDSnet, a national network of eight children's hospitals with data on >6.5 million children. Patients with three or more nephrologist encounters (n=55,560) not meeting the computable phenotype definition of glomerular disease were defined as nonglomerular cases. A reviewer blinded to case status used a standardized form to review random samples of cases (n=800) and nonglomerular cases (n=798). The final algorithm consisted of two or more diagnosis codes from a qualifying list or one diagnosis code and a pretransplant biopsy. Performance characteristics among the population with three or more nephrology encounters were sensitivity, 96% (95% CI, 94% to 97%); specificity, 93% (95% CI, 91% to 94%); positive predictive value (PPV), 89% (95% CI, 86% to 91%); negative predictive value, 97% (95% CI, 96% to 98%); and area under the receiver operating characteristics curve, 94% (95% CI, 93% to 95%). Requiring that the sum of nephrotic syndrome diagnosis codes exceed that of glomerulonephritis codes identified children with nephrotic syndrome or biopsy-based minimal change nephropathy, FSGS, or membranous nephropathy, with 94% sensitivity and 92% PPV. The algorithm identified 6657 children with glomerular disease across PEDSnet, ≥50% of whom were seen within 18 months. The authors developed an EHR-based algorithm and demonstrated that it had excellent classification accuracy across PEDSnet. This tool may enable faster identification of cohorts of pediatric patients with glomerular disease for observational or prospective studies.


The effects on pelvic visceral function of anal sphincter ablating and anal sphincter preserving operations for cancer of the lower part of the rectum and for benign colo-rectal disease.
The effects on pelvic visceral function of radical operations for carcinoma of the lower part of the rectum and of operations for inflammatory bowel disease have been determined by studying bladder, sexual and rectal function. Denervation of the bladder was diagnosed in 15% of patients who had undergone sphincter-saving resection of the rectum for carcinoma and in 19% of patients who had undergone abdominoperineal excision of the rectum. In the remaining patients, temporary, less severe, degrees of impairment of bladder function were noted. Abnormalities of function were found on urodynamic testing of patients with denervation of the bladder which explained, in part, the propensity of such patients to develop incontinence of urine. Partial recovery of function was noted in the long term in patients with denervation of the bladder and evidence consistent with regeneration of autonomic nerves was obtained on histological examination of biopsies from the bladders of such patients. In patients who had undergone conventional proctectomy for inflammatory bowel disease, impairment of bladder and sexual function was observed. In contrast, no evidence of damage to pelvic autonomic nerves was found in patients who underwent selective mucosal proctectomy. Mucosal proctectomy and ileo-anal anastomosis in patients with ulcerative colitis did impair ano-rectal reflex function; nevertheless, most patients were continent. Patients in whom, in addition to mucosal proctectomy and ileo-anal anastomosis, a pelvic ileal reservoir had been constructed had better rectal function than patients who lacked such a reservoir.


Intracellular antibody capture technology: application to selection of intracellular antibodies recognising the BCR-ABL oncogenic protein.
The expression of antibodies inside cells to ablate protein function has the potential for disease therapy and for target validation in functional genomics. However, due to inefficient expression or folding, only a few antibodies or antibody fragments, usually as single-chain Fv antibody fragments (scFv), bind their antigens in an intracellular environment. We have established a genetic-selection technology (intracellular antibody capture, IAC) to facilitate the isolation of functional intracellular scFv from a diverse repertoire. This approach comprises an in vitro library screen with scFv-expressing bacteriophage, employing bacterially expressed antigen, followed by a yeast in vivo antibody-antigen interaction screen of the sub-library of in vitro scFv antigen-binders. Accordingly, we have isolated panels of scFv that bind intracellularly to the BCR or the ABL parts of the BCR-ABL oncogenic protein. Sequence analysis of the intracellular antibody scFv panels revealed a sequence conservation indicating an intracellular antibody consensus for both VH and VL, which could form the basis for the de novo synthesis of intracellular antibody libraries to be used with intracellular antibody-capture technology.


Fumarate hydratase deficiency in renal cancer induces glycolytic addiction and hypoxia-inducible transcription factor 1alpha stabilization by glucose-dependent generation of reactive oxygen species.
Hereditary leiomyomatosis and renal cell cancer (HLRCC) is an inherited cancer syndrome linked to biallelic inactivation of the gene encoding the tricarboxylic acid cycle enzyme fumarate hydratase (FH). Individuals with HLRCC are at risk to develop cutaneous and uterine leiomyomas and an aggressive form of kidney cancer. Pseudohypoxic drive-the aberrant activation of cellular hypoxia response pathways despite normal oxygen tension-is considered to be a likely mechanism underlying the etiology of this tumor. Pseudohypoxia requires the oxygen-independent stabilization of the alpha subunit of the hypoxia-inducible transcription factor (HIF-1alpha). Under normoxic conditions, proline hydroxylation of HIF-1alpha permits VHL recognition and subsequent targeting for proteasomal degradation. Here, we demonstrate that inactivating mutations of FH in an HLRCC-derived cell line result in glucose-mediated generation of cellular reactive oxygen species (ROS) and ROS-dependent HIF-1alpha stabilization. Additionally, we demonstrate that stable knockdown of FH in immortalized renal epithelial cells results in ROS-dependent HIF-1alpha stabilization. These data reveal that the obligate glycolytic switch present in HLRCC is critical to HIF stabilization via ROS generation.


Modeling mental health information preferences during the early adult years: a discrete choice conjoint experiment.
Although most young adults with mood and anxiety disorders do not seek treatment, those who are better informed about mental health problems are more likely to use services. The authors used conjoint analysis to model strategies for providing information about anxiety and depression to young adults. Participants (N = 1,035) completed 17 choice tasks presenting combinations of 15 four-level attributes of a mental health information strategy. Latent class analysis yielded 3 segments. The virtual segment (28.7%) preferred working independently on the Internet to obtain information recommended by young adults who had experienced anxiety or depression. Self-assessment options and links to service providers were more important to this segment. Conventional participants (30.1%) preferred books or pamphlets recommended by a doctor, endorsed by mental health professionals, and used with a doctor's support. They would devote more time to information acquisition but were less likely to use Internet social networking options. Brief sources of information were more important to the low interest segment (41.2%). All segments preferred information about alternative ways to reduce anxiety or depression rather than psychological approaches or medication. Maximizing the use of information requires active and passive approaches delivered through old-media (e.g., books) and new-media (e.g., Internet) channels.


Trauma and adult attention deficit hyperactivity disorder.
This study investigated the relationship between adult attention deficit hyperactivity disorder (ADHD) and trauma. Fifty-eight adults admitted to hospital with musculoskeletal trauma were evaluated using scales that determine the presence of ADHD in childhood and adulthood. Each patient was also interviewed by an adult psychiatrist and a child and adolescent psychiatrist. The control group consisted of 30 adult patients with complaints other than trauma who did not have a history of repetitive traumas. There were 36 (62.2%) cases of ADHD in the patient group compared with four (13.3%) in the control group; this difference was statistically significant. When the level of trauma was evaluated, ADHD was identified in 23 of the 26 (88.5%) patients with high energy traumas compared with 14 of the 32 (43.8%) patients with low energy traumas; this difference was also statistically significant. This study shows that patients with adult ADHD are more prone to injuries, particularly high energy traumas such as motor vehicle accidents. Patients who have repeated high energy traumas should be evaluated by a psychiatrist for ADHD.


The loss of the BH3-only Bcl-2 family member Bid delays T-cell leukemogenesis in Atm-/- mice.
Multicellular organisms maintain genomic integrity and resist tumorigenesis through a tightly regulated DNA damage response (DDR) that prevents propagation of deleterious mutations either through DNA repair or programmed cell death. An impaired DDR leads to tumorigenesis that is accelerated when programmed cell death is prevented. Loss of the ATM (ataxia telangiectasia mutated)-mediated DDR in mice results in T-cell leukemia driven by accumulation of DNA damage accrued during normal T-cell development. Pro-apoptotic BH3-only Bid is a substrate of Atm, and Bid phosphorylation is required for proper cell cycle checkpoint control and regulation of hematopoietic function. In this report, we demonstrate that, surprisingly, loss of Bid increases the latency of leukemogenesis in Atm-/- mice. Bid-/-Atm-/- mice display impaired checkpoint control and increased cell death of DN3 thymocytes. Loss of Bid thus inhibits T-cell tumorigenesis by increasing clearance of damaged cells, and preventing propagation of deleterious mutations.


Verification of Prognostic Algorithms to Predict Remaining Flying Time for Electric Unmanned Vehicles.
This paper addresses the problem of building trust in the online prediction of a battery powered UAV's remaining available flying time. A series of ground tests is described that make use of an electric unmanned aerial vehicle (eUAV) to verify the performance of remaining flying time predictions. The algorithm verification procedure described is implemented on a fully functional vehicle that is restrained to a platform for repeated run-to-functional-failure (charge depletion) experiments. The vehicle under test is commanded to follow a predefined propeller RPM profile in order to create battery demand profiles similar to those expected during flight. The eUAV is repeatedly operated until the charge stored in powertrain batteries falls below a specified limit threshold. The time at which the limit threshold on battery charge is crossed is then used to measure the accuracy of the remaining flying time prediction. In our earlier work battery aging was not included. In this work we take into account aging of the batteries where the parameters were updated to make predictions. Accuracy requirements are considered for an alarm that warns operators when remaining flying time is estimated to fall below the specified limit threshold.


The individual and collective effects of exact exchange and dispersion interactions on the ab initio structure of liquid water.
In this work, we report the results of a series of density functional theory (DFT) based ab initio molecular dynamics (AIMD) simulations of ambient liquid water using a hierarchy of exchange-correlation (XC) functionals to investigate the individual and collective effects of exact exchange (Exx), via the PBE0 hybrid functional, non-local van der Waals/dispersion (vdW) interactions, via a fully self-consistent density-dependent dispersion correction, and an approximate treatment of nuclear quantum effects, via a 30 K increase in the simulation temperature, on the microscopic structure of liquid water. Based on these AIMD simulations, we found that the collective inclusion of Exx and vdW as resulting from a large-scale AIMD simulation of (H2O)128 significantly softens the structure of ambient liquid water and yields an oxygen-oxygen structure factor, SOO(Q), and corresponding oxygen-oxygen radial distribution function, gOO(r), that are now in quantitative agreement with the best available experimental data. This level of agreement between simulation and experiment demonstrated herein originates from an increase in the relative population of water molecules in the interstitial region between the first and second coordination shells, a collective reorganization in the liquid phase which is facilitated by a weakening of the hydrogen bond strength by the use of a hybrid XC functional, coupled with a relative stabilization of the resultant disordered liquid water configurations by the inclusion of non-local vdW/dispersion interactions. This increasingly more accurate description of the underlying hydrogen bond network in liquid water also yields higher-order correlation functions, such as the oxygen-oxygen-oxygen triplet angular distribution, POOO(θ), and therefore the degree of local tetrahedrality, as well as electrostatic properties, such as the effective molecular dipole moment, that are in much better agreement with experiment.


Landscape-level spatial patterns of West Nile virus risk in the northern Great Plains.
Understanding the landscape-level determinants of West Nile virus (WNV) can aid in mapping high-risk areas and enhance disease control and prevention efforts. This study analyzed the spatial patterns of human WNV cases in three areas in South Dakota during 2003-2007 and investigated the influences of land cover, hydrology, soils, irrigation, and elevation by using case-control models. Land cover, hydrology, soils, and elevation all influenced WNV risk, although the main drivers were different in each study area. Risk for WNV was generally higher in areas with rural land cover than in developed areas, and higher close to wetlands or soils with a high ponding frequency. In western South Dakota, WNV risk also decreased with increasing elevation and was higher in forested areas. Our results showed that the spatial patterns of human WNV risk were associated with landscape-level features that likely reflect variability in mosquito ecology, avian host communities, and human activity.


Nurse case-manager vs multifaceted intervention to improve quality of osteoporosis care after wrist fracture: randomized controlled pilot study.
Few outpatients with fractures are treated for osteoporosis in the years following fracture. In a randomized pilot study, we found a nurse case-manager could double rates of osteoporosis testing and treatment compared with a proven efficacious quality improvement strategy directed at patients and physicians (57% vs 28% rates of appropriate care). Few patients with fractures are treated for osteoporosis. An intervention directed at wrist fracture patients (education) and physicians (guidelines, reminders) tripled osteoporosis treatment rates compared to controls (22% vs 7% within 6 months of fracture). More effective strategies are needed. We undertook a pilot study that compared a nurse case-manager to the multifaceted intervention using a randomized trial design. The case-manager counseled patients, arranged bone mineral density (BMD) tests, and prescribed treatments. We included controls from our first trial who remained untreated for osteoporosis 1-year post-fracture. Primary outcome was bisphosphonate treatment and secondary outcomes were BMD testing, appropriate care (BMD test-treatment if bone mass low), and costs. Forty six patients untreated 1-year after wrist fracture were randomized to case-manager (n = 21) or multifaceted intervention (n = 25). Median age was 60 years and 68% were female. Six months post-randomization, 9 (43%) case-managed patients were treated with bisphosphonates compared with 3 (12%) multifaceted intervention patients (relative risk [RR] 3.6, 95% confidence intervals [CI] 1.1-11.5, p = 0.019). Case-managed patients were more likely than multifaceted intervention patients to undergo BMD tests (81% vs 52%, RR 1.6, 95%CI 1.1-2.4, p = 0.042) and receive appropriate care (57% vs 28%, RR 2.0, 95%CI 1.0-4.2, p = 0.048). Case-management cost was $44 (CDN) per patient vs $12 for the multifaceted intervention. A nurse case-manager substantially increased rates of appropriate testing and treatment for osteoporosis in patients at high-risk of future fracture when compared with a multifaceted quality improvement intervention aimed at patients and physicians. Even with case-management, nearly half of patients did not receive appropriate care. clinicaltrials.gov identifier: NCT00152321.


Interactions between membrane conductances underlying thalamocortical slow-wave oscillations.
Neurons of the central nervous system display a broad spectrum of intrinsic electrophysiological properties that are absent in the traditional "integrate-and-fire" model. A network of neurons with these properties interacting through synaptic receptors with many time scales can produce complex patterns of activity that cannot be intuitively predicted. Computational methods, tightly linked to experimental data, provide insights into the dynamics of neural networks. We review this approach for the case of bursting neurons of the thalamus, with a focus on thalamic and thalamocortical slow-wave oscillations. At the single-cell level, intrinsic bursting or oscillations can be explained by interactions between calcium- and voltage-dependent channels. At the network level, the genesis of oscillations, their initiation, propagation, termination, and large-scale synchrony can be explained by interactions between neurons with a variety of intrinsic cellular properties through different types of synaptic receptors. These interactions can be altered by neuromodulators, which can dramatically shift the large-scale behavior of the network, and can also be disrupted in many ways, resulting in pathological patterns of activity, such as seizures. We suggest a coherent framework that accounts for a large body of experimental data at the ion-channel, single-cell, and network levels. This framework suggests physiological roles for the highly synchronized oscillations of slow-wave sleep.


Linear Response Equilibrium versus echo-planar encoding for fast high-spatial resolution 3D chemical shift imaging.
In this work Linear Response Equilibrium (LRE) and Echo-planar spectroscopic imaging (EPSI) are compared in terms of sensitivity per unit time and power deposition. In addition an extended dual repetition time scheme to generate broad stopbands for improved inherent water suppression in LRE is presented. The feasibility of LRE and EPSI for assessing cholesterol esters in human carotid plaques with high spatial resolution of 1.95×1.15×1.15 mm(3) on a clinical 3T MR system is demonstrated. In simulations and phantom experiments it is shown that LRE has comparable but lower sensitivity per unit time relative to EPSI despite stronger signal generated. This relates to the lower sampling efficiency in LRE relative to EPSI as a result of limited gradient performance on clinical MR systems. At the same time, power deposition of LRE is significantly reduced compared to EPSI making it an interesting niche application for in vivo high field spectroscopic imaging of metabolites within a limited bandwidth.


A Computational Synaptic Antibody Characterization Tool for Array Tomography.
Application-specific validation of antibodies is a critical prerequisite for their successful use. Here we introduce an automated framework for characterization and screening of antibodies against synaptic molecules for high-resolution immunofluorescence array tomography (AT). The proposed Synaptic Antibody Characterization Tool (SACT) is designed to provide an automatic, robust, flexible, and efficient tool for antibody characterization at scale. SACT automatically detects puncta of immunofluorescence labeling from candidate antibodies and determines whether a punctum belongs to a synapse. The molecular composition and size of the target synapses expected to contain the antigen is determined by the user, based on biological knowledge. Operationally, the presence of a synapse is defined by the colocalization or adjacency of the candidate antibody punctum to one or more reference antibody puncta. The outputs of SACT are automatically computed measurements such as target synapse density and target specificity ratio that reflect the sensitivity and specificity of immunolabeling with a given candidate antibody. These measurements provide an objective way to characterize and compare the performance of different antibodies against the same target, and can be used to objectively select the antibodies best suited for AT and potentially for other immunolabeling applications.


The practice of Burch Colposuspension versus Mid Urethral Slings for the treatment of Stress Urinary Incontinence in developing country.
To compare the effectiveness and complications of Burch colposuspension and Mid Urethral Slings (MUS) for the treatment of Stress Urinary Incontinence (SUI). We conducted a cross-sectional study of 162 patients who underwent surgery for SUI with Burch colposuspension (n=40), tension free vaginal tape (TVT) (n= 59) or transobturator tape (TOT) (n=63), from 2006 to 2014 at the Aga Khan University Hospital- Karachi. All three groups were assessed in terms of demographics, cure rates, intraoperative and postoperative complications at one and five years using incontinence impact questionnaire-short form-7 (IIQ-7) and urogenital distress inventory -short form-6 (UDI-6). Mean age of the participants in Burch, TVT and TOT group was 44.1 ± 7.4, 48.3 ± 8.9, 53.0 ± 9.4 respectively. Majority of patients in TVT group were premenopausal (59.3%) and postmenopausal in TOT group (53.9%). Most abdominal hysterectomies were done in Burch group (40) while vaginal hysterectomies and anterior and posterior colporrhaphy in TOT group (55). All the procedures had both subjective and objective cure rate of more than 82% at one year, with TVT having the highest success rate of 96.61%. The objective cure rate in Burch, TVT and TOT group at five years was 74.19%, 90.30% and 81.25% respectively. Intraoperative complications included hemorrhage in one patient during Burch procedure and bladder perforation in two cases of TVT, with no significant difference in short or long-term complications with either procedure. All the three procedures have equal efficacy and complication rates. Even though TVT is the new gold standard but in view of current debate regarding mesh related complications, there is a need to readdress Burch colposuspension for treatment of SUI.


Mitochondrial dysfunction causing cardiac sodium channel downregulation in cardiomyopathy.
Cardiomyopathy is associated with cardiac Na(+) channel downregulation that may contribute to arrhythmias. Previously, we have shown that elevated intracellular NADH causes a decrease in cardiac Na(+) current (I(Na)) signaled by an increase in mitochondrial reactive oxygen species (ROS). In this study, we tested whether the NADH-mitochondria ROS pathway was involved in the reduction of I(Na) in a nonischemic cardiomyopathic model and correlated the findings with myopathic human hearts. Nonischemic cardiomyopathy was induced in C57BL/6 mice by hypertension after unilateral nephrectomy, deoxycorticosterone acetate (DOCA) pellet implantation, and salt water substitution. Sham operated mice were used as controls. After six weeks, heart tissue and ventricular myocytes isolated from mice were utilized for whole cell patch clamp recording, NADH/NAD(+) level measurements, and mitochondrial ROS monitoring with confocal microscopy. Human explanted hearts were studied using optical mapping. Compared to the sham mice, the arterial blood pressure was higher, the left ventricular volume was significantly enlarged (104.7±3.9 vs. 87.9±6.1 μL, P<0.05), and the ejection fraction was reduced (37.1±1.8% vs. 49.4±3.7%, P<0.05) in DOCA mice. Both the whole cell and cytosolic NADH level were increased (279±70% and 123±2% of sham, respectively, P<0.01), I(Na) was decreased (60±10% of sham, P<0.01), and mitochondrial ROS overproduction was observed (2.9±0.3-fold of sham, P<0.01) in heart tissue and myocytes of myopathic mice vs. sham. Treatment of myocytes with NAD(+) (500 μM), mitoTEMPO (10 μM), chelerythrine (50 μM), or forskolin (5 μM) restored I(Na) back to the level of sham. Injection of NAD(+) (100mg/kg) or mitoTEMPO (0.7 mg/kg) twice (at 24h and 1h before myocyte isolation) to animals also restored I(Na). All treatments simultaneously reduced mitochondrial ROS levels to that of controls. CD38 was found to transduce the extracellular NAD(+) signal. Correlating with the mouse model, failing human hearts showed a reduction in conduction velocity that improved with NAD(+). Nonischemic cardiomyopathy was associated with elevated NADH level, PKC activation, mitochondrial ROS overproduction, and a concomitant decrease in I(Na). Reducing mitochondrial ROS by application of NAD(+), mitoTEMPO, PKC inhibitors, or PKA activators, restored I(Na). NAD(+) improved conduction velocity in human myopathic hearts.


Cerebellar ataxia induced by 3-AP affects immunological function.
We previously showed that the cerebellum modulates the immune system. Here we determined whether cerebellar ataxia alters immunological function to further demonstrate an involvement of the cerebellum in immune modulation. Neurotoxin 3-acetylpyridine (3-AP) was intraperitoneally injected in rats to induce cerebellar ataxia. Behavior and motor coordination were tested on day 7 following 3-AP injection. Nissl staining and high-performance liquid chromatography (HPLC) were used to determine neuronal loss and neurotransmitter contents, respectively, in all the three cerebellar nuclei, fastigial nucleus (FN), interposed nucleus (IN) and dentate nucleus (DN). T and B lymphocyte differentiation and function were measured by flow cytometry, Western blot and ELISA. 3-AP induced motor discoordination and locomotor reduction. In all the three cerebellar nuclei, FN, IN and DN, there was a neuronal loss and a decrease in contents of glutamate and GABA (but not glycine) after 3-AP injection. Importantly, CD4+ T cells, but not CD8+ T cells, were increased by the 3-AP treatment. Moreover, interferon (IFN)-γ-producing cells and interleukin (IL)-17-producing cells were decreased in cerebellar ataxia rats, but IL-4-producing cells and CD25-expressing cells were increased. Expression of the T helper (Th)1- and Th17-related cytokines, IFN-γ, IL-2, IL-17 and IL-22, was downregulated in CD4+ cells in cerebellar ataxia rats, while expression of the Th2 and regulatory T (Treg)-related cytokines, IL-4, IL-5, IL-10 and transforming growth factor (TGF)-β, was upregulated. Furthermore, B lymphocyte number and anti-bovine serum albumin (BSA) IgM and IgG antibody levels were elevated in cerebellar ataxia. Cerebellar ataxia alters cellular and humoral immunity.